ApoNifH functions in iron–molybdenum cofactor synthesis and apodinitrogenase maturation

Abstract

NifH (dinitrogenase reductase) has three important roles in the nitrogenase enzyme system. In addition to its role as the obligate electron donor to dinitrogenase, NifH is required for the iron-molybdenum cofactor (FeMo-co) synthesis and apodinitrogenase maturation. We have investigated the requirement of the Fe-S cluster of NifH for these processes by preparing apoNifH. The 4Fe-4S cluster of NifH was removed by chelation of the cluster with ␣, ␣-bipyridyl. The resulting apoNifH was tested in in vitro FeMo-co synthesis and apodinitrogenase maturation reactions and was found to function in both these processes. Thus, the presence of a redox active 4Fe-4S cluster in NifH is not required for its function in FeMo-co synthesis and in apodinitrogenase maturation. This, in turn, implies that the role of NifH in these processes is not one of electron transfer or of iron or sulfur donation.

In Vitro Biosynthesis of Iron-Molybdenum Cofactor and Maturation of the nif-encoded Apodinitrogenase

“…The ability of these forms of NifH, impaired in catalysis and with altered MgATP reactivities, to function in FeMo-co synthesis and apodinitrogenase maturation strongly suggests a role(s) for NifH independent of its role in substrate reduction. We have reported previously that the presence of a redox-active 4Fe-4S cluster in NifH is not required for its function in FeMo-co synthesis and in apodinitrogenase maturation (35). This is further evidence in support of distinct roles for NifH in nitrogenase turnover, FeMo-co biosynthesis, and apodinitrogenase maturation.…”

Biosynthesis of the Iron-Molybdenum Cofactor of Nitrogenase

“…An alternative interpretation is that the impairment of nifUS mutants in the P-cluster assembly is in fact reflecting a deficient assembly of the [Fe 4 -S 4 ] cluster of NifH. We do not favor this alternative interpretation because there is evidence showing that [Fe 4 -S 4 ] cluster-deficient NifH functions in apo-NifDK maturation (14,61).…”

Accumulation of 55Fe-Labeled Precursors of the Iron-Molybdenum Cofactor of Nitrogenase on NifH and NifX ofAzotobacter vinelandii

“…It has also been known that the properties of NifH required for its function in FeMo-co biosynthesis are different from those required for its function in electron transfer to dinitrogenase (34 -36). For example, we have shown in a separate study that a redox-active 4Fe-4S cluster in NifH is not necessary for its function in FeMo-co biosynthesis (37). Nevertheless, the exact role played by NifH in cofactor biosynthesis remains unclear.…”